Functional analysis of the endothelial cell-specific Tie2/Tek promoter identifies unique protein-binding elements.
To investigate the molecular basis of endothelial cell-specific gene expression, we have examined the DNA sequences and the cognate DNA-binding proteins that mediate transcription of the murine tie2/tek gene. Reporter transfection experiments conformed with earlier findings in transgenic mice, indicating that the upstream promoter of Tie2/Tek is capable of activating transcription in an endothelial cell-specific fashion. These experiments have also allowed the identification of a single upstream inhibitory region (region I) and two positive regulatory regions (regions U and A) in the proximal promoter. Electrophoretic mobility-shift assays have allowed further characterization of three novel DNA-binding sequences associated with these regions and have provided preliminary characterization of the protein factors binding to these elements. Two of the elements (U and A) confer increased transcription on a heterologous promoter, with element U functioning in an endothelial-cell-selective manner. By employing embryonic endothelial-like yolk sac cells in parallel with adult-derived endothelial cells, we have identified differences in functional activity and protein binding that may reflect mechanisms for specifying developmental regulation of tie2/tek expression. Further study of the DNA and protein elements characterized in these experiments is likely to provide new insight into the molecular basis of developmental- and cell-specific gene expression in the endothelium.
Fadel
B M
BM
Division of Cardiology, Vanderbilt University Medical Center, 315 MRB II, Nashville, TN 37232-6300, USA.
Boutet
S C
SC
Quertermous
T
T
D000818:Animals; D001483:Base Sequence; D002417:Cattle; D018983:DNA Footprinting; D004268:DNA-Binding Proteins / Q000378:metabolism; D004730:Endothelium, Vascular / Q000502:physiology*; D005786:Gene Expression Regulation; D051379:Mice; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D051381:Rats; D020794:Receptor Protein-Tyrosine Kinases / Q000235:genetics*; D042787:Receptor, TIE-2; D015183:Restriction Mapping; D014158:Transcription, Genetic; D014162:Transfection
D004268:DNA-Binding Proteins; D020794:Receptor Protein-Tyrosine Kinases; D042787:Receptor, TIE-2